Chemical Synthesis

The clickable chelators were obtained starting from the tetrazine containing precursors Tz (5) and Tz-PEG (6). These precursors were successively coupled with the commercially available DOTA-NHS ester and DOTAGA-anhydride, as shown in Figure 1. The couplings were carried out in dry DMF in the presence of NEt₃ and by stirring the reaction mixture at room temperatures during 3 h to afford the desired final compounds DOTA-Tz (1), DOTAPEG-Tz (2), DOTAGA-Tz (3) and DOTAGAPEG-Tz (4). The chemical synthesis of the derivative DOTAGA-Tz (3) has been previously reported (23). The clickable TCO-containing bombesin antagonist (TCO-PEG4-AR) was performed by coupling the peptide D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH₂ (AR) with TCO-PEG4-NHS, in solution and in presence of DIPEA. The chemical synthesis, purification and characterization of the different compounds are detailed in the Supplementary Material.